System and method for in vitro analysis of therapeutic agents

ABSTRACT

A system and method for in vitro analysis of therapeutic agents comprising a reservoir adapted to hold a therapeutic agent, a first flow cell having a first cell chamber adapted to receive at least a first sample of the therapeutic agent, a second flow cell having a second cell chamber adapted to receive at least a second sample of the therapeutic agent, the first flow cell having a first path length (b e ′) and the second flow cell having a second path length (b e ″), the first path length being substantially equal to a sensitivity factor (f)×b e ″, a membrane chamber having a biological cell membrane therein adapted to receive at least a third sample of the therapeutic agent, the membrane chamber being further adapted to detect the membrane potential of the biological cell membrane; and spectroscopic detection means for detecting the spectral characteristics of the first and second therapeutic agent samples.

This application claims the benefit of 60/264,363, filed Jan. 26, 2001.

FIELD OF THE PRESENT INVENTION

The present invention relates generally to electrophysiologic assessmentof therapeutic agents. More particularly, the invention relates to asystem and method for in vitro assessment of therapeutic agents thatemploys spectroscopic means for accurate determination of the agent'sconcentration.

BACKGROUND OF THE INVENTION

The potential of cardiovascular and non-cardiovascular therapeuticagents or drugs to cause prolongation of the QT (i.e., cardiacrepolarization time between two ventricular sequences) interval of theelectrocardiogram has been, and continues to be, a significant factor inthe development of new therapeutic agents. Indeed, it is wellestablished that a wide range of non-cardiovascular therapeutic agentsthat are not expected on the basis of their mechanism of action toprolong QT can produce a substantial number of serious cardiac events.Such agents belong to different pharmacological classes, such aspsychotropic drugs (e.g., tricyclic-amitriptiline and tetracyclicantidepressants, phenothiazine derivatives, haloperidol, pimozide,risperidone and sertindole), prokinetic (e.g., cisapride), antimalarialmedicines (e.g., halofantrine, quinine, and chloroquine), antibioticsbelonging to several chemical classes (e.g., azithromycin, erythromycin,clarithromycin, spiramycin, pentamidine, trimethoprim-sulfamethoxazoleand sparfloxacin), antifungal agents (e.g., ketoconazole, fuconazole anditraconazole), agents for treating urinary incontinence (e.g.,terodiline), and certain histamine H₁-receptor antagonist (e.g.,astemizole, terfenadine and diphenhydramine).

These therapeutic agents, in certain very rare instances, can triggerlife-threatening polymorphic ventricular tachycardias, such as torsadede pointes, often in the presence of additional factors favoring,directly or indirectly, proarrhythmic events. The relevant factorsinclude congenital or acquired long-QT syndrome, ischemic heart disease,congestive heart failure, severe hepatic or renal dysfunction,bradycardia, electrolyte imbalance (e.g., hypokalemia due to diuretictreatment, hypomagnesemia, hypocalcemia, acidosis and intracellular Ca⁺⁺loading), intentional or accidental overdose, and concomitant treatmentwith ion channel blocking drugs or agents that inhibit the drugdetoxification processes.

Several preclinical techniques have thus been employed to evaluate thecardiovascular effects of proposed therapeutic agents. The notedtechniques to determine concentration of proposed therapeutic agentsprimarily comprise high performance liquid chromatography (HPLC) orother analytical assays that are generally limited to higher therapeuticagent concentrations unless pre-concentration or large volumes areemployed.

It is, however, well known that the noted physiological analyticalassays are often laborious, expensive, time consuming and frustrated bytechnical problems. Further, HPLC and/or assays may require hours todays to analyze samples and process data depending on the complexity andnumber of samples.

It is therefore an object of the present invention to provide a systemand method for high-speed, economical in vitro analyses of therapeuticagents.

It is another object of the present invention to provide a system andmethod for in vitro analysis of low concentration therapeutic agents.

It is yet another object of the present invention to provide a systemand method for correlating the electrophysiological effects of aproposed therapeutic agent over a broad concentration range.

SUMMARY OF THE INVENTION

In accordance with the above objects and those that will be mentionedand will become apparent below, in a preferred embodiment, the systemfor in vitro analysis of therapeutic agents in accordance with thisinvention comprises a reservoir adapted to hold a therapeutic agent, afirst flow cell having a first cell chamber adapted to receive at leasta first sample of the therapeutic agent, a second flow cell having asecond cell chamber adapted to receive at least a second sample of thetherapeutic agent, the first flow cell having a first path length(b_(e)′) and the second flow cell having a second path length (b_(e)″)the first path length being substantially equal to a sensitivity factor(f)×b_(e)″, a membrane chamber having a biological cell membrane thereinadapted to receive at least a third sample of the therapeutic agent, themembrane chamber being further adapted to detect the membrane potentialof the biological cell membrane, and spectroscopic detection means fordetecting the spectral characteristics of the first and secondtherapeutic agent samples.

The method for in vitro analysis of therapeutic agents in accordancewith the invention preferably comprises the steps of (i) introducing afirst sample of a therapeutic agent into a first flow cell having afirst path length (b_(e)′), (ii) introducing a second sample of thetherapeutic agent into a second flow cell having a second path length(b_(e)″), the first path length (b_(e)′) being substantially equal to asensitivity factor (f)×b_(e)″, (iii) introducing a third sample of thetherapeutic agent into membrane chamber means having a biological cellmembrane disposed therein, (iv) measuring the absorption spectrum of thefirst therapeutic agent sample by transmitting a given wavelength of afirst light into the first flow cell, (v) measuring the absorptionspectrum of the second therapeutic agent sample by transmitting a givenwavelength of a second light into the second flow cell, and (vi)detecting the membrane potential of said biological cell membrane.

BRIEF DESCRIPTION OF THE DRAWINGS

Further features and advantages will become apparent from the followingand more particular description of the preferred embodiments of theinvention, as illustrated in the accompanying drawings, and in whichlike referenced characters generally refer to the same parts or elementsthroughout the views, and in which:

FIG. 1 is a schematic illustration of one embodiment of the in vitrospectroscopic system for analysis of therapeutic agents according to theinvention;

FIG. 2 is a partial section plan view of one embodiment of a flow cellaccording to the invention;

FIG. 3 is a partial section plan view of an additional embodiment of aflow cell according to the invention;

FIG. 4 is a schematic illustration of a flow cell body showing theattenuated light path according to the invention;

FIG. 5 is a graphical illustration of absorbance spectra of apharmaceutical composition having a low concentration therapeutic agentand a high concentration therapeutic agent;

FIG. 6 is a schematic illustration of the spectroscopic means accordingto the invention; and

FIG. 7 is a partial section perspective view of the membrane chambermeans according to the invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The method and system of the present invention substantially reduces oreliminates the drawbacks and shortcomings associated with prior artelectrophysiologic assessment of therapeutic agents. As discussed indetail below, the system generally includes a plurality of flow cells,spectroscopic means in communication with the flow cells, membranechamber means and flow passage means for introducing the subjecttherapeutic agent(s) to the flow cells and membrane chamber means.

By the term “therapeutic agent”, as used herein, it is meant to mean andinclude active ingredients, components or elements of a pharmaceuticalcomposition, drugs and medicaments.

Cardiac action potential (or membrane potential) is generally defined asthe pattern of electrical activity that is associated with excitablebiological cells (e.g., heart cells). It is the result of numerous,distinct, successively activated currents generated by the passage ofbiologically important ions (Na⁺, Ca⁺⁺ and K⁺) through specializedmembrane structures such as ionic pumps and exchangers and, mostimportantly, voltage-gated ion channels. These currents are consideredto be depolarizing when they carry extracellular positive charges intothe cell and to be repolarizing when they carry positive charges to thecell exterior.

Therapeutic agents that modify the normal flux of ions through channelscan, and in many instances will, modify certain aspects of the membranepotential and, thus, affect cardiac function. Indeed, blockers of Na⁺channels reduce the rate of rise of the membrane potential (Vmax) andcan produce disturbances in cardiac conduction, which, if severe, may belife threatening. Agents that decrease the rate of Na-currentinactivation and increase residual Na-current can prolong the durationof the action potential (ADP), prolong the QT interval and thus maytrigger torsades de pointes arrhythmias. Blockers of Ca⁺⁺ channelsgenerally decrease ADP, reduce the rate of A-V conduction and producecardiac depression, whereas Ca⁺⁺ channel-activators prolong ADP and maycause arrhythmias. Finally, K⁺ channel-blockers prolong ADP and QT andcan provoke arrhythmias, whereas K⁺ channel-activators shorten ADP andcan also trigger arrhythmia.

Thus, an effective indicator (or parameter) of possible adverse cardiaceffects of a proposed therapeutic agent is the change in membranepotential (i.e., membrane resting potential) resulting from theintroduction or exposure to the therapeutic agent. Indeed, the notedparameter is often deferred to in conventional physiological assessmentsof proposed therapeutic agents (or drug candidates). As discussed indetail below, the noted parameter is also employed in the presentinvention to assess the potential physiological effect(s) of atherapeutic agent.

Referring now to FIG. 1, there is shown a schematic illustration of oneembodiment of the in vitro analysis system 5 of the invention. Asillustrated in FIG. 1, the system 5 preferably includes reservoir means(e.g. reservoir) 10, optic based spectroscopic detection means 20, aplurality of flow cells, preferably first and second flow cells 60, 70,membrane chamber means 40 and flow passage means 12.

According to the invention, the reservoir means 10 is designed andadapted to store at least one therapeutic agent in diluent. As will beappreciated by one having ordinary skill in the art, various capacity(and configuration) reservoir means 10 may be employed within the scopeof the invention. In a preferred embodiment of the invention, thecapacity of the reservoir means 10 is in the range of 1 to 2000 ml.

The reservoir means 10 is further designed and adapted to receive theflow passage means 12 of the invention, which preferably comprisessubstantially non-adsorbing tubing (e.g., stainless steel, PEEK®. Asillustrated in FIG. 1, pump means 14 is also provided to facilitate flowof the therapeutic agent (i.e., therapeutic agent sample or samples)from the reservoir means 10 to and through the first and second flowcells 60, 70. The pump means 14 is in communication with the flowpassage means 12 and is preferably disposed between the reservoir means10 and the flow cells 60, 70. According to the invention, the pump means14 is capable of achieving a sample flow rate in the range of ≦0.5 to≧10 ml/min., more preferably, in the operating range of approximately 2to 5 ml/min.

As will be appreciated by one having ordinary skill in the art, variouspump means 14 may be employed within the scope of the invention toprovide the noted sample flow rate(s). In a preferred embodiment, thepump means 14 comprises a peristaltic pump.

Referring now to FIG. 2, there is shown one embodiment the first flowcell 60 of the invention. For simplicity, only the first flow cell 60will be illustrated and described. However, it is to be understood thatthe second flow cell 70 of the noted embodiment is similarly constructedand the description of the first flow cell 60 is equally applicable toeach flow cell 60, 70.

As illustrated in FIG. 2, the flow cell 60 preferably includes asubstantially tubular body 62 having an inlet port 64, an outlet port 66and a cell chamber, designated generally 63, disposed therein that isadapted to receive a therapeutic agent sample. According to theinvention, the inlet and outlet ports 64, 66 and cell chamber 63 definea flow passage 61.

As will be appreciated by one having ordinary skill in the art, variousflow cell body 62 materials may be employed within the scope of theinvention. In a preferred embodiment of the invention, the body 62 ofeach flow cell 60, 70 includes a core comprising a polymer, silica,chalcogenide or other like materials and cladding 67 disposed on theouter surface of the core comprising a polymer or doped silica or otherlike materials (see FIGS. 2 and 4).

According to the invention, the first flow cell 60 further includesfirst light transmission means 21 a adapted to provide a givenwavelength of excitation light or radiation (and/or a given rangethereof) to the flowable therapeutic agent present in the first flowcell chamber 63 (i.e., first sample) and first light detection means 23a for detecting the transmitted light from the first sample. The secondflow cell 70 similarly includes second light transmission means 21 badapted to provide a given wavelength of excitation light (and/or agiven range thereof) to the sample present in the cell chamber of thesecond flow cell 70 (i.e., second sample) and second light detectionmeans 23 b for detecting the transmitted light from the second sample.

Referring now to FIG. 3, there is shown another embodiment of a flowcell 80 of the invention. The flow cell 80 similarly includes an inletport 82, an outlet port 84 and a cell chamber, designated generally 86,disposed there between that is similarly adapted to receive atherapeutic agent sample.

The flow cell 80 further includes first light transmission means 85adapted to provide a given wavelength of excitation light or radiationto the flowable therapeutic agent present in the flow cell chamber 86(i.e., first sample) and first light detection means 87 for detectingthe transmitted light from the first sample.

It is well known that when each sample passes through a respective flowcell chamber (e.g., 63) the amount of excitation light transmitted intoand through the cell chamber 63 decreases in accordance with Beer's Law,i.e.,

$\begin{matrix}{A = {\frac{I}{I_{o}} = 10^{- {\propto {b\; c}}}}} & {{Eq}.\mspace{14mu} 1}\end{matrix}$where:

-   

-   A=absorbance;

-   I=power of transmitted radiation;

-   I_(o)=power of incident radiation;

-   c=molar absorptivity of the sample;

-   c=sample concentration (moles/liter); and

-   b=path length of the light in the chamber (cm.)

The absorbance (A) is thus defined as the product of ∝bc. According toBeer's Law, absorbance (A) is also proportional to both the sampleconcentration (c) and path length (b).

As will be appreciated by one having ordinary skill in the art, the pathlength (b) is generally deemed a “straight path length” that isapplicable in light transmission/detection configurations such as thatillustrated in FIG. 3. It will further be appreciated by one havingskill in the art that the noted Beer's Law relationship is similarlyapplicable for the light transmission/detection configuration (i.e., 21a, 23 a) of the cylindrical flow cell 60 of the invention (see FIG. 2),which employs attenuated total reflection.

Referring to FIG. 4, it is well known that when radiation (or light),designated generally 100, undergoes total internal reflection 102, itactually penetrates a fraction of a wavelength into the medium (orsample) beyond the reflecting surface. The penetration depth isgenerally denoted d_(p).

Since the penetration depth d_(p) is defined as a unitlength/reflection, an equivalent path length (b_(e)) can thus be derivedas follows:b _(e) =d _(p) ×R×L  Eq.2where:

-   d_(p)=penetration depth per reflection;-   R=number of reflections per unit length; and-   L=length of tube or cell.

The equivalent path length (b_(e)) can then be employed in Eq.1 toderive the absorbance of a respective sample (A).

As will further be appreciated by one having ordinary skill in the art,the effective path length (b_(e)) of a respective flow cell 60, 70 isdirectly dependent on the length of the flow cell body 62. Thus, thelength of each flow cell 60, 70 can be tailored to provide differentpath lengths.

It can further be deduced from Beer's Law that if the absorbance rangeof the spectroscopic means (e.g., spectrophotometer) is fixed, which isa common element of conventional spectroscopic means, the path length (bor b_(e)) must be increased for lower molar absorptivity (∝) or lowerconcentration (c). However, as is well known in the art, the responseband generally associated with a larger path length is undesirablynarrow and, hence, limited.

It is also well known in the art that a broader response band can beachieved by employing two (2) flow cells having different path lengths(b_(e)′, b_(e)″). The typical path lengths generally range from 0.1 to100 cm.

Applicants have however found that an optimum dynamic (i.e.,substantially linear) response range can be achieved if b_(e)′ issubstantially equal to b_(e)″ ×a sensitivity factor (f). According tothe invention, the sensitivity factor (f) preferably has a value in therange of 1 to 100. More preferably, f has a value in the range of 1 to20.

Referring to FIG. 5, there are shown absorbance spectra of a lowconcentration therapeutic agent (i.e., ˜18 ng/ml) and a highconcentration therapeutic agent (i.e., ˜3800 ng/ml). The notedabsorbance spectra was derived with a first flow cell having a pathlength of approx. 5 cm and a second flow cell having a path cell ofapprox. 50 cm (i.e., f=10).

Referring to Curve A, it can be seen that a path length of 5 cm wasinsufficient to detect the low concentration therapeutic agent over arange of wavelengths from 200 nm to 400 nm. However, as illustrated byCurve B, the same agent was readily detectable (and quantifiable) at apath length of 50 cm.

Referring now to Curve C, it can be seen that the high concentrationtherapeutic agent was not quantifiable at a path length of 50 cm.However, as illustrated in Curve D, the same agent was readilydetectable at a path length of 5 cm.

Accordingly, in a preferred embodiment of the invention, the first flowcell 60 has an effective path length (b_(e)′) in the range of 0.1 to 100cm and second flow cell 70 has an effective path length (b_(e)″) in therange of 0.1 to 100 cm. More preferably, the first flow cell 60 has aneffective path length (b_(e)′) in the range of approx. 5 to 50 cm andthe second flow cell 70 has a path length (b_(e)″) in the range ofapprox. 50 to 100 cm. Applicants have found that accurate spectralcharacteristics of therapeutic agents having a substantially lowconcentration in the range of 5 to 10 ng/ml can readily be detected byvirtue of the noted range of path lengths (b_(e)′, b_(e)″).

Referring now to FIG. 6, in addition to the first and second lighttransmission means 21 a, 21 b and first and second light detection means23 a, 23 b shown in FIG. 2, the spectroscopic means 20 of the inventionfurther includes light source means 22 for providing the desiredwavelength of light to the first and second light transmission means 21a, 21 b via optical lines 24 a and 24 b, respectively, and analyzermeans 26 for analyzing the light detected by the first and second lightdetection means 23 a, 23 b, which is communicated to the analyzer means26 via optical lines 28 a and 28 b, respectively.

In additional envisioned embodiments of the invention, the reservoirmeans 10 also includes light transmission means 25 a and light detectionmeans 25 b that are operatively connected to the spectroscopic means 20of the invention via optical lines 27 a, 27 b. As will be appreciated byone having ordinary skill in the art, the reservoir means lighttransmission and detection means 25 a, 25 b provides means forsimultaneously assessing and monitoring the agent contained in thereservoir means 10 and, hence, means for detecting therapeutic agentloss and ensuring that the samples analyzed in the flow cells 60, 70 arerepresentative of the source therapeutic agent contained in thereservoir means 10.

As illustrated in FIG. 6, the spectroscopic means 20 further preferablyincludes memory means 30 for storing at least the detected spectroscopiccharacteristics of the first and second samples (and source agentcontained in the reservoir means 10) and the control parameters for thespectroscopic means of the invention, processor means 32 for processingat least the spectroscopic characteristics of the samples (and sourceagent) and first display means 34 (shown in phantom) for displaying atleast the spectroscopic characteristics of the samples, the sourcepharmaceutical agent contained in the reservoir means 10 and the“processed” spectroscopic characteristics of the samples (e.g., meanvalues).

As will be appreciated by one having ordinary skill in the art, variousconventional light source means 22 and/or analyzer means 26 can beemployed within the scope of the invention to provide a given range ofwavelength of light, analyze the spectroscopic characteristics (e.g.,absorption spectrum of the absorbed light) acquired by the first andsecond light detection means 23 a, 23 b and control the spectroscopicmeans 20 of the invention, such as the analyzers disclosed in U.S. Pat.No. 4,664,522 and the MCS-521 fiber optic UV/VIS spectrophotometerdistributed by Carl Zeiss, which are incorporated by reference herein.The analyzer means 26 and/or the processor means 32 may also comprise apersonal computer.

Referring now to FIG. 7, there is shown the membrane chamber means 40 ofthe invention. The membrane chamber means 40 includes a membrane chamberbody 42 having a perfusion inlet 44, a perfusion outlet 46, apreparation well 48, a membrane well 50 and a diffuser plate 52 disposedbetween the preparation well 48 and membrane well 50.

As illustrated in FIG. 7, the membrane chamber means 40 further includesmembrane means 52 disposed in the membrane well 50. By the term“membrane means”, as used herein, it is meant to mean a biological cellmembrane, including a sheet or layer of a biological organ, theventricular muscle and/or papillary muscle, and Purkinje fibers.

In a preferred embodiment of the invention, the membrane means 52comprises a Purkinje fiber. As is well known in the art, Purkinje fibersare often employed in electrophysiological assessments because it isbelieved that the major ionic currents underlying their actionpotentials resemble those contributing to the repolarization process ofthe human heart.

To assess the membrane potential of the membrane means 52, the membranechamber means 40 further includes a plurality of electrodes. Referringto FIG. 7, at least one, preferably two, bipolar electrodes 54 a, 54 bare operatively connected to the membrane means 52 proximate one endthereof. The electrodes 54 a, 54 b are also in communication with thestimulation means 55 of the invention via leads 54 a, 54 b (see FIG. 1)and are adapted to provide a stimulating charge or current to themembrane means 52.

As illustrated in FIG. 7, a further electrode is also provided. In apreferred embodiment, the noted electrode comprises an intracellularmicroelectrode 56 that is also operatively connected to the membranemeans 52.

According to the invention, the microelectrode 56 is designed andadapted to detect the membrane potential of the membrane means 52. Themicroelectrode 56 is preferably in communication with second displaymeans 58 (via lead 57 c) that is adapted to provide a visual display orindication of the detected potential (see FIG. 1).

Assessment of therapeutic agents in accordance with the presentinvention is preferably accomplished as follows: The spectroscopic meansof the invention is initially calibrated by conventional means. Suchmeans includes analysis of blank (or primary) samples as a UV referenceand an analysis of calibration samples with respective blank samples asa reference.

After calibration of the spectroscopic means 20, flow of the therapeuticagent (preferably, in diluent) is initiated and the therapeutic agent isintroduced into and through the flow passage means 12 via pump means 14.The therapeutic agent is then introduced into and through the flowpassage 61 of the first and second flow cells 60, 70, which arepreferably connected in series, and the membrane chamber means 40.

The spectroscopic characteristics of the therapeutic agent present inthe first flow cell 60 (i.e., first sample) and the second flow cell 70(i.e., second sample) are then detected (preferably, substantiallysimultaneously) by the above described spectroscopic means 20 of theinvention. The spectroscopic characteristics are then processed andanalyzed by conventional means.

In a preferred embodiment, substantially simultaneously with thespectroscopic analysis and while the therapeutic agent is present in themembrane well 50 of the membrane chamber means 40 (i.e., third sample),the membrane means 52 is subjected to an initial current via stimulatingelectrodes 54 a, 54 b. The membrane potential is then detected viaelectrode 56 that is displayed on the second display means 58 of theinvention. The change in membrane potential is then readily determinedby comparing the detected membrane potential to the potential of themembrane means 52 prior to exposure to the therapeutic agent.

As will be appreciated by one having ordinary skill in the art, thesystem and method of the invention, described in detail above, is alsoapplicable to analyses of pharmaceutical compositions containing atherapeutic agent and, in particular, pharmaceutical compositions havinga substantially low concentration of therapeutic agents.

SUMMARY

From the foregoing, one of ordinary skill in the art can easilyascertain that the present invention provides the following advantages:

1. High speed, highly accurate and economical (i.e., low cost) in vitroanalyses of therapeutic agents.

2. High speed, efficient in vitro analyses of pharmaceuticalcompositions having a substantially low concentration (i.e., 5–10 ng/ml)of therapeutic agents.

3. High speed, efficient means for correlating the electrophysiologicaleffects of a proposed therapeutic agent over a broad concentrationrange.

4. Means for assessing and monitoring the stability of the therapeuticagent during in vitro analyses.

5. Means for assessing and monitoring agent loss (e.g., glasswareabsorption and/or attachment) during in vitro analyses.

6. Means for rapidly and efficiently detecting carryover (i.e.,cross-contamination) in a reservoir and/or feed lines.

Without departing from the spirit and scope of this invention, one ofordinary skill can make various changes and modifications to theinvention to adapt it to various usages and conditions. As such, thesechanges and modifications are properly, equitably, and intended to be,within the full range of equivalence of the following claims.

1. A system for use in in-vitro analysis of a therapeutic agent, saidsystem comprising: a reservoir adapted to hold said therapeutic agent; afirst flow cell in communication with said reservoir having a first cellchamber adapted to receive at least a first sample of said therapeuticagent; a second flow cell in communication with said reservoir having asecond cell chamber adapted to receive at least a second sample of saidtherapeutic agent; said first flow cell having a first path length(b_(e)′) and said second flow cell having a second path length (b_(e)″),said first path length being substantially equal to sensitivity factor(f)×b_(e)″; a membrane chamber in communication with said reservoirhaving a biological cell membrane therein, said membrane chamber beingadapted to receive at least a third sample of said therapeutic agent,said membrane chamber being further adapted to detect the membranepotential of said biological cell membrane; and spectroscopic detectionmeans for detecting the spectral characteristics of said first andsecond therapeutic agent samples.
 2. The system of claim 1, wherein saidspectroscopic detection means includes first light transmission meansfor transmitting a first light of a given wavelength into said firstcell chamber, first light detection means for detecting a firsttransmitted light from said first therapeutic agent sample, second lighttransmission means for transmitting a second light of a given wavelengthinto said second cell chamber and second light detection means fordetecting a second transmitted light from said second therapeutic agentsample.
 3. The system of claim 2, wherein said spectroscopic detectionmeans further includes control means in communication with said firstand second light transmission means and said first and second lightdetection means for providing said first and second lights and analyzingsaid first and second transmitted lights.
 4. The system of claim 1,wherein said sensitivity factor has a value in the range of 1 to
 100. 5.The system of claim 4, wherein said sensitivity factor has a value inthe range of 1 to
 20. 6. The system of claim 1, wherein said first andsecond path lengths are in the range of 0.1 to 100 cm.
 7. The system ofclaim 6, wherein said first path length is in the range of 5 to 50 cm.8. The system of claim 6, wherein said second path length is in therange of 50 to 100 cm.
 9. The system of claim 1, wherein saidspectroscopic detection means includes third light transmission meansfor transmitting a third light of a given wavelength into said reservoirand third light detection means for detecting a third transmitted lightfrom said therapeutic agent contained in said reservoir, said thirdlight transmission means and third detection means being incommunication with said control means.
 10. The system of claim 1,wherein said spectroscopic means includes first display means fordisplaying at least the spectroscopic characteristics of said first andsecond therapeutic agent samples.
 11. The system of claim 1, whereinsaid membrane chamber means includes second display means for displayingsaid membrane potential of said biological cell membrane.
 12. A systemfor use in in-vitro analysis of a therapeutic agent, said systemcomprising: a reservoir adapted to receive said therapeutic agent; afirst flow cell in communication with said reservoir having a first cellchamber adapted to receive a first sample of said therapeutic agent,said first flow cell having an effective path length in the range ofapproximately 5–50 cm; a second flow cell in communication with saidreservoir having a second cell chamber adapted to receive a secondsample of said therapeutic agent, said second flow cell having aneffective path length in the range of approximately 50–100 cm; amembrane chamber in communication with said reservoir having abiological cell membrane therein, said membrane chamber being adapted toreceive a third sample of said therapeutic agent, said membrane chamberbeing further adapted to detect the membrane potential of saidbiological cell membrane; and spectroscopic detection means fordetecting the spectral characteristics of said first and secondtherapeutic agent samples.
 13. The system of claim 12, wherein saidspectroscopic detection means includes first light transmission meansfor transmitting a first light of a given wavelength into said firstcell chamber, first light detection means for detecting a firsttransmitted light from said first therapeutic agent sample, second lighttransmission means for transmitting a second light of a given wavelengthinto said second cell chamber and second light detection means fordetecting a second transmitted light from said second therapeutic agentsample.
 14. The system of claim 13, wherein said spectroscopic detectionmeans further includes control means in communication with said firstand second light transmission means and said first and second lightdetection means for providing said first and second lights and analyzingsaid first and second transmitted lights.
 15. The system of claim 14,wherein said spectroscopic detection means includes third lighttransmission means for transmitting a third light of a given wavelengthinto said reservoir and third light detection means for detecting athird transmitted light from said therapeutic agent contained in saidreservoir, said third light transmission means and third detection meansbeing in communication with said control means.
 16. The system of claim15, wherein said spectroscopic means includes display means fordisplaying at least the spectroscopic characteristics of saidtherapeutic agent contained in said reservoir and said first and secondtherapeutic agent samples.
 17. A method for in-vitro analysis oftherapeutic agents, said method comprising the steps of: introducing atleast a first sample of a therapeutic agent into a first flow cell, saidfirst flow cell having a first path length (b_(e)′); introducing atleast a second sample of said therapeutic agent into a second flow cell,said second flow cell having a second path length (b_(e)″); said firstpath length (b_(e)′) being substantially equal to a sensitivity factor(f)×b_(e)″; introducing at least a third sample of said therapeuticagent into membrane chamber means having a biological cell membranedisposed therein; measuring the absorption spectrum of said firsttherapeutic agent sample by transmitting a given wavelength of a firstlight into said first flow cell; measuring the absorption spectrum ofsaid second therapeutic agent sample by transmitting a given wavelengthof a second light into said second flow cell; and detecting the membranepotential of said biological cell membrane.
 18. The method of claim 17,wherein said first therapeutic agent sample absorption spectrum and saidsecond therapeutic agent sample absorption spectrum are measuredsubstantially simultaneously.
 19. The method of claim 17, wherein saidsensitivity factor has a value in the range of 1 to
 100. 20. The methodof claim 17, wherein said sensitivity factor has a value in the range of1 to
 20. 21. The method of claim 17, wherein said first and second pathlengths are in the range of 0.1 to
 100. 22. The system of claim 21,wherein said first path length is in the range of 5 to 50 cm.
 23. Thesystem of claim 21, wherein said second path length is in the range of50 to 100 cm.